We will continue biochemical experiments concerning the synthesis and processing of cell surface glycoproteins. The glycoproteins of animal cells are extracellular proteins which comprise most of the protein of the cell surface, the extracellular matrix and extracellular fluids. The mechanism of secretion of glycoproteins and its coupling to their synthesis is a problem that has attracted much attention in recent years. Palade and coworkers showed that secretory proteins are made on membrane-bound ribosomes in the rough endoplasmic reticulum and then transported via the golgi to the cell surface. The mechanism by which peptides cross the endoplasmic reticulum membrane has also been elucidated by Blobel, Sabbatini and others. Chains begun on soluble ribosomes attach to the membrane by a short N-terminal "signal sequence"; the nascent chain is then extruded across the membrane, so that the finished product is released directly into the lumen. It is also known that carbohydrate residues are added both in the endoplasmic reticulum and the golgi. However, it is not known when or in what chemical form sugar residues cross the membrane. We will approach this problem by determining the transmembrane localization of lipid-linked oligosaccharide, the central intermediate in the synthesis of asparagine-linked oligosaccharides. We will also continue experiments on the glycosylation pattern of recombinant RNA viruses. Finally, we will continue enzymatic studies on the mechanisms of assembly of the dolichol pyrophosphate linked oligosaccharide.